PROJECT SUMMARY Acute immune responses involving CD8+ cytotoxic T lymphocytes (CTLs) and/or natural killer cells can effectively restrain tumor development and progression. Unfortunately, immunotherapy attempts to date have struggled to achieve significant clinical benefit as single agents in PC. This is likely due to the presence of an immunosuppressive tumor microenvironment. Critical drivers of this immunosuppressive microenvironment are tumor-infiltrating inflammatory monocytes (IMs) and macrophages (TAMs). Thus, high numbers of these cells correlate with early metastatic relapse and poor survival in pancreatic cancer. Therefore, approaches that reprogram myeloid responses to potentiate protective antitumor immunity hold significant therapeutic potential. Targeting tumor-infiltrating myeloid cells to improve therapeutic outcomes: We and other groups have demonstrated that mobilization and tumor infiltration of IMs and TAMs can promote local immunosuppression, and resistance to cytotoxic therapy. Signaling through C-C chemokine receptor type 2 (CCR2) is critical for the mobilization of IMs and their recruitment to inflamed tissues. Our recently published reports clearly illustrate that blockade of IM recruitment using a novel CCR2 inhibitor, PF-04136309 (CCR2i), slows tumor progression, improves responses to chemotherapy and prevents metastasis in mouse models of PC1,13. Based on these exciting and provocative data, we initiated a Phase Ib/II clinical trial targeting the CCR2 signaling pathway in patients with locally advanced PC. In this trial, we have observed a remarkable 48.5% response rate in the 33 patients treated with CCR2i + FOLFIRINOX. Additionally, this regimen was well tolerated (safe). These responses appear to be correlated with a marked reduction in circulating CCR2+ IMs as well as decreased immune suppressive gene expression profiles in the primary tumor microenvironment. Paralleling these clinical data, our published pre-clinical studies found that CCR2 blockade overcomes immune suppression to reinitiate anti-tumor responses via CD8+ CTLs. Intriguingly, we've discovered that CCR2 blockade in both human patients and mouse models leads to the upregulation of T cell checkpoint pathways, including programmed cell death-1 (PD1) and its ligands. These data suggest that we might find unique therapeutic synergy between CCR2 inhibition and PD1-based immunotherapies. Thus, we propose the following aims: Aim 1: Determine the effects of CCR2 blockade on T lymphocyte responses in patients with PC. Aim 2: Determine the mechanisms by which CCR2 inhibition improves T cell immunity. Aim 3: Determine whether combining CCR2 inhibition with PD1 blockade can enhance therapeutic outcomes in patients with advanced PC. Summary: The proposed research will assess the safety and efficacy of targeting CCR2 to improve PD1 based immunotherapy. At the same time, we will improve our understanding of the mechanism(s) by which CCR2 blockade improves CTL responses in humans and mice.